Dries A M Feyen1,2, Isaac Perea-Gil1,3, Renee G C Maas4, Magdalena Harakalova4, Alexandra A Gavidia3, Jennifer Arthur Ataam1,3, Ting-Hsuan Wu3, Aryan Vink5, Jiayi Pei4, Nirmal Vadgama1,3, Albert J Suurmeijer6, Wouter P Te Rijdt7,8, Michelle Vu1,2, Prashila L Amatya1,2, Maricela Prado3, Yuan Zhang3, Logan Dunkenberger3, Joost P G Sluijter4, Karim Sallam1,2, Folkert W Asselbergs4,9,10,11, Mark Mercola1,2, Ioannis Karakikes1,3. 1. Cardiovascular Institute, Stanford University, CA (D.A.M.F., I.P.-G., J.A.A., N.V., M.V., P.L.A., K.S., M.M., I.K.). 2. Departments of Medicine (D.A.M.F., M.V., P.L.A., K.S., M.M.), Stanford University School of Medicine, Stanford University, CA. 3. Cardiothoracic Surgery (I.P.-G., A.A.G., J.A.A., T.-H.W., N.V., M.P., Y.Z., L.D., I.K.), Stanford University School of Medicine, Stanford University, CA. 4. Department of Cardiology, Division Heart and Lungs (R.G.C.M., M.H., J.P., J.P.G.S., F.W.A.), University Medical Center Utrecht, University of Utrecht, The Netherlands. 5. Department of Pathology (A.V.), University Medical Center Utrecht, University of Utrecht, The Netherlands. 6. Department of Pathology, University Medical Center Groningen, University of Groningen, The Netherlands (A.J.S.). 7. Netherlands Heart Institute, Utrecht (W.P.t.R.). 8. Department of Genetics, University Medical Center Groningen, University of Groningen, The Netherlands (W.P.t.R.). 9. Institute of Cardiovascular Science, Faculty of Population Health Sciences (F.W.A.), University College London, UK. 10. Health Data Research UK (F.W.A.), University College London, UK. 11. Institute of Health Informatics (F.W.A.), University College London, UK.
Abstract
BACKGROUND: Phospholamban (PLN) is a critical regulator of calcium cycling and contractility in the heart. The loss of arginine at position 14 in PLN (R14del) is associated with dilated cardiomyopathy with a high prevalence of ventricular arrhythmias. How the R14 deletion causes dilated cardiomyopathy is poorly understood, and there are no disease-specific therapies. METHODS: We used single-cell RNA sequencing to uncover PLN R14del disease mechanisms in human induced pluripotent stem cells (hiPSC-CMs). We used both 2-dimensional and 3-dimensional functional contractility assays to evaluate the impact of modulating disease-relevant pathways in PLN R14del hiPSC-CMs. RESULTS: Modeling of the PLN R14del cardiomyopathy with isogenic pairs of hiPSC-CMs recapitulated the contractile deficit associated with the disease in vitro. Single-cell RNA sequencing revealed the induction of the unfolded protein response (UPR) pathway in PLN R14del compared with isogenic control hiPSC-CMs. The activation of UPR was also evident in the hearts from PLN R14del patients. Silencing of each of the 3 main UPR signaling branches (IRE1, ATF6, or PERK) by siRNA exacerbated the contractile dysfunction of PLN R14del hiPSC-CMs. We explored the therapeutic potential of activating the UPR with a small molecule activator, BiP (binding immunoglobulin protein) inducer X. PLN R14del hiPSC-CMs treated with BiP protein inducer X showed a dose-dependent amelioration of the contractility deficit in both 2-dimensional cultures and 3-dimensional engineered heart tissues without affecting calcium homeostasis. CONCLUSIONS: Together, these findings suggest that the UPR exerts a protective effect in the setting of PLN R14del cardiomyopathy and that modulation of the UPR might be exploited therapeutically.
BACKGROUND: Phospholamban (PLN) is a critical regulator of calcium cycling and contractility in the heart. The loss of arginine at position 14 in PLN (R14del) is associated with dilated cardiomyopathy with a high prevalence of ventricular arrhythmias. How the R14 deletion causes dilated cardiomyopathy is poorly understood, and there are no disease-specific therapies. METHODS: We used single-cell RNA sequencing to uncover PLN R14del disease mechanisms in human induced pluripotent stem cells (hiPSC-CMs). We used both 2-dimensional and 3-dimensional functional contractility assays to evaluate the impact of modulating disease-relevant pathways in PLN R14del hiPSC-CMs. RESULTS: Modeling of the PLN R14del cardiomyopathy with isogenic pairs of hiPSC-CMs recapitulated the contractile deficit associated with the disease in vitro. Single-cell RNA sequencing revealed the induction of the unfolded protein response (UPR) pathway in PLN R14del compared with isogenic control hiPSC-CMs. The activation of UPR was also evident in the hearts from PLN R14del patients. Silencing of each of the 3 main UPR signaling branches (IRE1, ATF6, or PERK) by siRNA exacerbated the contractile dysfunction of PLN R14del hiPSC-CMs. We explored the therapeutic potential of activating the UPR with a small molecule activator, BiP (binding immunoglobulin protein) inducer X. PLN R14del hiPSC-CMs treated with BiP protein inducer X showed a dose-dependent amelioration of the contractility deficit in both 2-dimensional cultures and 3-dimensional engineered heart tissues without affecting calcium homeostasis. CONCLUSIONS: Together, these findings suggest that the UPR exerts a protective effect in the setting of PLN R14del cardiomyopathy and that modulation of the UPR might be exploited therapeutically.
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